Overview of therapeutic strategies

Though AR appears to be an inevitable consequence of EGFR TKI treatment, successful treatment strategies to overcome AR have been elusive. More than 75 studies have looked at over 50 investigational drugs and drug combinations in the setting of AR to first-generation EGFR TKI therapy, with the majority of studies reporting little to no efficacy in this population (Table 2). In this review, we will discuss completed and ongoing efforts to overcome resistance.

Published or presented studies with results in patients with EGFR-mutant lung cancer and AR to EGFR TKI

Therapeutic Strategies Targeting EGFR

Standard treatment with addition of continued EGFR inhibition beyond progression

Management of patients with progression on first-line EGFR TKI therapy is largely dependent on clinical factors such as symptoms and disease burden. Some patients may have RECIST progression based on tumor measurements, but with continued clinical benefit from therapy. In this setting, many asymptomatic patients can delay a change in therapy for several months to over a year (17). This approach is being studied prospectively in the ASPIRATION trial (18). If there is limited progression, local therapy and continued EGFR inhibition can be considered (19, 20). For patients with oligometastatic disease, a prolonged PFS after local therapy of 6 to 10 months was achieved in several small series (19, 20).

There are limited prospective data to support continued EGFR TKI in addition to chemotherapy in the AR setting. Erlotinib has been combined with platinum doublet chemotherapy with toxicity that is similar to chemotherapy alone (4). In a retrospective series of 78 patients with EGFR-mutant lung cancer and AR, the OR for response rate (RR) was 0.20 favoring chemotherapy with concurrent erlotinib compared with chemotherapy alone (21). In a trial comparing afatinib and paclitaxel to chemotherapy alone in patients with disease progression on single-agent EGFR TKI, there was an increase in PFS with continuation of EGFR TKI (5.6 vs. 2.8 months, HR 0.60; ref. 22). There are several ongoing prospective studies that may result in a definitive recommendation about the combination of chemotherapy and erlotinib (NCT01928160, NCT02064491, and NCT02098954) or gefitinib (NCT01544179) in the AR setting.

Second-generation EGFR TKIs

Second-generation EGFR TKIs bind to the EGFR tyrosine kinase domain irreversibly and have broader ERBB inhibition (Table 1). Afatinib is a dual EGFR/HER2 inhibitor that is now FDA approved for the first-line treatment of lung cancers with EGFR L858R mutations or exon 19 deletions. This approval is based on a phase III trial that demonstrated a superior PFS for afatinib compared with cisplatin and pemetrexed (13.6 vs. 6.9 months) in patients with EGFR-mutant lung cancer (3).

Although afatinib has proven useful in the first-line setting, its role in AR is less clear. The LUX-Lung 1 trial randomized molecularly unselected patients, who had previously been treated with erlotinib or gefitinib for at least 3 months, to receive afatinib or placebo (23). The RR (7% vs. <1%) and PFS (3.3 vs. 1.1 months) were superior with afatinib compared with placebo, but there was no improvement in overall survival (the primary endpoint) in all study participants or in the subset of patients with known EGFR-mutant lung cancer.

Neratinib has activity against both EGFR and HER2 and was evaluated in a phase II study in patients with non–small cell lung cancer (NSCLC; ref. 24). The RR was 2% among those patients with EGFR mutations, but there were no responses in patients with EGFR T790M. Dacomitinib, a pan-HER inhibitor, was administered to patients with NSCLC whose disease had progressed on chemotherapy and erlotinib (25). In the subset of patients with EGFR-mutant lung cancer, the RR was 8%; no responses were seen in patients with EGFR T790M. The dual EGFR/VEGFR inhibitors, XL647 and vandetanib, have also been tested in the AR setting with limited efficacy observed (26, 27).

EGFR antibodies

Cetuximab, an EGFR monoclonal antibody, binds to the extracellular domain of EGFR and prevents ligand-dependent receptor activation. Cetuximab has predominantly been used in combination therapy with EGFR TKIs in the resistance setting. In a phase I/II study of erlotinib plus cetuximab in patients with AR to erlotinib, no responses (0/19 evaluable patients) were seen (28). However, the combination of cetuximab and afatinib has shown more promise (29). In a clinical trial of afatinib and cetuximab in 100 patients with AR to erlotinib or gefitinib, treatment with the combination led to a 30% overall RR and a median PFS of 4.7 months (30). Responses were seen in patients with both EGFR T790M-positive and T790M-negative tumors. The most common toxicities were rash, diarrhea, and fatigue. Randomized trials of afatinib plus cetuximab are planned in both the first-line and AR settings.

Mutant-specific EGFR inhibitors

A relatively new class of drugs irreversibly inhibits mutant EGFR, in particular EGFR T790M, with much less activity against wild-type EGFR (Table 1). The first drug in this class, WZ4002, is 30- to 100-fold more potent against EGFR T790M and up to 100-fold less potent against wild-type EGFR in preclinical models (31). This discovery led to the subsequent development of other compounds in this same class. AP26113 is a reversible dual ALK and EGFR inhibitor that has selectivity for the mutant forms of EGFR, both activating and resistance mutations, in preclinical models (32). The phase I/II of AP26113 demonstrated no objective responses in 12 patients with EGFR T790M containing tumors (33).

CO-1686 targets mutant EGFR, including T790M, and is active in EGFR-mutant xenografts and murine models of erlotinib resistance (34). The phase I study of single-agent CO-1686 is complete with a small range of doses being tested in the ongoing phase II study (35). CO-1686 has the notable adverse event of hyperglycemia as a frequent adverse event (38%, any grade). The reported overall RR is 58% in 40 patients with centrally confirmed EGFR T790M tumors. Median PFS has not been reached, but is estimated at >12 months (35).

AZD9291 is another mutant specific EGFR TKI currently being tested clinically (36). A phase I study of AZD9291 in patients with advanced NSCLC who had progression on first- or second-generation EGFR TKI therapy is nearing completion (37). As of April 2014, 232 patients had received AZD9291 with no dose-limiting toxicities identified. Diarrhea (14%, any grade) and rash (24%, any grade) were the most frequent toxicities. The RR in 107 patients with EGFR T790M tumors to date was 64%.

Several other EGFR-mutant specific TKIs are being tested in phase I studies currently, including HM61713 (NCT01588145), EGF816 (NCT02108964), and ASP8273 (NCT02113813). Preliminary results with HM61713 were recently presented (38). One-hundred and eighteen patients have received the drug, and an RR of 22% was noted in the 83 evaluable patients. The RR was 29% in 48 patients with EGFR T790M containing tumors. EGF816 similarly has selective activity against mutant EGFR in cellular assays and xenograft models (39).

Therapeutic Strategies Targeting Alternate Pathways

MET inhibition

MET is a receptor tyrosine kinase which, when activated by its ligand, hepatocyte growth factor (HGF), can mediate cell proliferation, evasion of apoptosis, and metastasis (40). MET amplification is seen in untreated patients with NSCLC at a rate of approximately 4% (41, 42), and is detected in approximately 5% of tumors with AR to EGFR TKI (5, 6). De novoMET amplification has been associated with primary resistance to EGFR TKIs (43). Therapeutic targeting of MET may be an effective strategy in MET-amplified tumors (44, 45). Results from a phase II study of cabozantinib (an oral MET/VEGFR2/RET inhibitor) in patients with EGFR-mutant lung cancer and progression on EGFR TKI resulted in three partial responses out of 35 patients treated (46). There are several ongoing clinical trials of MET TKIs or MET MAbs in combination with EGFR TKIs with the majority of studies selecting for patients that are MET positive by various assays (Table 3). An ongoing phase I study of INC280 and gefitinib in EGFR-mutant patients with AR who have either MET amplification or MET overexpression has modest activity, with a 15% unconfirmed RR (6/41 patients; ref. 47). In addition to MET amplification, MET activation through increased production of HGF is a potential mechanism of resistance to EGFR TKIs (48). Lung cancer cell lines made resistant to EGFR TKIs by HGF overexpression were sensitive to dual EGFR and MET blockade (49). Prospective clinical validation of anti–HGF-directed monoclonal antibodies is pending.

Ongoing and recently completed studies without presented or published results in patients with EGFR-mutant lung cancers

HER2 and HER3 inhibition

The ErbB family of receptor tyrosine kinases is composed of four members—EGFR (ErbB1), HER2 (ErbB2), HER3 (ErbB3), and HER4 (ErbB4). HER2 amplification has been detected in 12% of tumors with AR to EGFR TKI therapy (13), resulting in sustained downstream signaling, even in the continued presence of the EGFR TKI. Combination strategies involving dual EGFR/HER2 blockade are being explored. The combination of the dual HER2/EGFR TKI, afatinib plus cetuximab, has been studied in patients with AR to erlotinib (30), as described above. In a phase I/II clinical trial of erlotinib plus the HER3 monoclonal antibody, MM-121 (NCT00994123), one EGFR TKI naïve patient had a partial response and five of eight patients with EGFR-mutant/TKI-resistant disease had stable disease (50).

IGF-IR inhibition

The insulin like growth factor-1 receptor (IGF-IR) is a receptor tyrosine kinase that is activated by the IGF-I or IGF-II ligands (51). Upregulation of IGF-IR signaling has previously been described to mediate AR to first-generation EGFR TKIs (52) as well as to the irreversible EGFR inhibitors PF299804 and WZ4002 (53). Though IGF-IR was implicated in the pathogenesis of EGFR-mutant lung cancer, clinical testing of combined EGFR/IGF-IR blockade has not proven to be effective. A randomized phase II study of erlotinib alone or in combination with the IGF-IR/Insulin receptor TKI, OSI-906, in patients with advanced EGFR-mutant NSCLC (NCT01221077) closed early after an interim analysis in March 2013 showed that there would be no benefit to the combination.

AXL inhibition

Activation of the receptor tyrosine kinase AXL has been reported as a mechanism of resistance to erlotinib. Recent studies have demonstrated increased AXL activation, without EGFR T790M mutation, in both in vitro and in vivo models of EGFR-mutant/TKI-resistant lung cancer (10). Combined inhibition of AXL and EGFR in these models restored therapeutic efficacy. More recently, increased AXL expression was observed in 5 of 26 patients (19%) with AR to gefitinib (54). AXL inhibitors are in early clinical development (NCT00697632, ISRCTN00759419; ref. 55).

FGFR inhibition

More recently, in vitro studies have demonstrated increased expression of FGFR2 and its ligand, FGF1, in EGFR-mutant cell lines derived to be resistant to gefitinib or afatinib (14, 15). Combined inhibition of EGFR and FGFR in these models improved therapeutic responses in the EGFR TKI-resistant cell lines. Validation of these findings in clinical models has yet to be demonstrated.

MAPK pathway inhibition

Activation of the MAPK signaling pathway (Fig. 1) can drive EGFR TKI resistance in both preclinical models as well as in patient tumor samples. In a large series of tumor biopsy samples from patients with EGFR-mutant/TKI resistant lung cancer, 2 of 195 (1%) were found to have mutations in BRAF (9). Concomitant inhibition of EGFR and MEK was an effective therapeutic strategy in models of EGFR TKI resistance driven by MAPK pathway activation (9). Erlotinib resistance has also been associated with reduced expression of neurofibromin, a RAS GTPase-activating protein encoded by the NF1 gene which functions as a negative regulator of RAS (56). In preclinical models of EGFR TKI resistance, erlotinib failed to fully inhibit MAPK signaling in the context of low neurofibromin levels; however, these tumors responded to the combination of an EGFR TKI with a MEK inhibitor. Downregulation of NF1 expression was also seen in clinical samples from patients with EGFR TKI resistance. There are several ongoing clinical trials of MEK inhibitors plus EGFR TKIs in EGFR-mutant lung cancer (Table 3). In addition, one of the few described mechanisms of resistance to the mutant-specific EGFR TKIs is aberrant activation of ERK signaling (57). The MAPK pathway may become increasingly relevant as we develop combination strategies to prevent or overcome resistance to EGFR mutant-specific TKIs.

PI3K–AKT–mTOR pathway inhibition

Analogous to the MAPK pathway, activation of the PI3K–AKT–mTOR pathway (Fig. 1) also drives EGFR TKI resistance. PIK3CA mutations can co-occur with EGFR mutations and may portend a poorer response to EGFR TKIs (58–60). In addition, acquired PIK3CA mutations have been detected in a small percentage (∼5%) of EGFR-mutant lung cancers with AR to EGFR TKIs (6). Preclinical data have shown that introduction of activating PIK3CA mutations into EGFR-mutant cell lines confers resistance to EGFR TKIs (61). A phase II clinical trial combining erlotinib with the PI3K inhibitor, BKM120, is ongoing in patients with advanced NSCLC previously sensitive to erlotinib or whose tumors harbor an EGFR mutation (NCT01487265), although no PIK3CA mutation or aberration is required. A phase II study combining erlotinib with the AKT inhibitor, MK-2206, in patients with EGFR-mutant lung cancers and progression on erlotinib demonstrated a 9% RR (4/46 patients; ref. 62). Preclinical data also support the use of mTOR inhibitors in the context of EGFR TKI resistance (63), although a small study of EGFR TKI and everolimus in patients who previously responded to EGFR TKI demonstrated no responses (64). Concomitant inhibition of EGFR and mTOR in patients with AR is currently being evaluated further in a phase I trial of afatinib plus sirolimus (NCT00993499).

HSP90 inhibitors

Heat shock protein 90 (HSP90), a molecular chaperone for several oncogenic kinases, is required for protein folding and stabilization of mutant EGFR. Single-agent activity of the HSP90 inhibitor, AUY922, has been documented in patients with EGFR-mutant lung cancer, with a RR of 20% (7/35 patients; ref. 65). In a phase II study combining AUY922 with erlotinib, in patients with EGFR-mutant/TKI-resistant lung cancer, the overall RR was 2 of 16 (13%; ref. 66). Several HSP90 inhibitors are in clinical development for use in both EGFR TKI-sensitive and EGFR TKI-resistant patient populations (Table 3).

Immune Therapy

Another potential strategy that has garnered much attention recently is the use of immune checkpoint inhibitors, either as single agents or in combination with EGFR TKIs. An intriguing hypothesis is that tumor cell death triggered by targeted therapies results in antigen release and immunomodulation that can prime tumors and potentiate antitumor immune responses (67). Preclinical data in EGFR-mutant lung cancer cell lines and mouse models has demonstrated that mutant EGFR signaling drives expression of programmed death-ligand 1 (PD-L1), and that blockade of the PD-1 receptor improved survival of mice with EGFR-mutant tumors (68). A phase I clinical trial of erlotinib plus the anti–PD-L1 antibody, MPDL3280A, in the first-line setting (NCT02013219), a phase I/II study of pembrolizumab in combination with erlotinib or gefitinib (NCT02039674), and a study of erlotinib and nivolumab (NCT01454102) are all currently ongoing. Preliminary results indicate activity of the nivolumab and erlotinib combination; an overall RR of 19% (4/21 patients) was reported, with 3 of 4 responders having previously progressed on erlotinib monotherapy (69).

Future Directions

Despite the tremendous success of first-generation EGFR TKIs in patients with EGFR-mutant lung cancer, the emergence of resistance remains a significant barrier for the successful management of patients with this disease. Several strategies are beginning to show promise. In particular, continued clinical development of the new mutant specific EGFR TKIs has the potential to dramatically change our treatment paradigm for patients with EGFR-mutant lung cancers. However, AR that does not involve EGFR T790M is a heterogeneous clinical problem with more limited success.

Moving forward, there are several critical issues that remain to be addressed: (i) How clinically relevant is heterogeneity in resistance mechanisms to first-generation EGFR TKIS? Published reports have already documented co-occurrence of more than one resistance mechanism (e.g., EGFR T790M and MET amplification) within a given tumor sample, and among different sites of disease. (ii) As patients are treated with multiple lines of EGFR-targeted therapies, what are the mechanisms to mutant-specific EGFR inhibitors as sequential treatment? How does sequential treatment with multiple EGFR inhibitors affect tumor evolution and resistance? Will mutant-specific EGFR inhibitors make EGFR T790M rare? (iii) What are the optimal sequences of anti–EGFR-directed therapies and combination therapies? Will combination therapies and mutant-specific EGFR TKI be successful in the first-line setting? Will combination strategies, including combinations of small molecules inhibiting various targets or combinations of EGFR inhibitors with chemotherapy or immune therapy, overcome resistance? (iv) What are the best treatments for patient with lung cancers harboring noncanonical EGFR mutations, such as exon 20 insertions? (v) Are immune therapies useful in molecularly defined cohorts? As repeat biopsies increasingly become the standard of care, matching treatments targeting a specific resistance mechanism to patients whose tumors harbor that mechanism will allow us to more accurately assess therapeutic efficacy. Additional study of EGFR TKI resistance may provide clinicians with a greater understanding of therapeutic resistance across the numerous other cancers in which targeted therapies are being used.

Disclosure of Potential Conflicts of Interest

H.A. Yu is a consultant/advisory board member for Clovis Oncology. G.J. Riely reports receiving commercial research support from Novartis and Roche, and is a consultant/advisory board member for Novartis. C.M. Lovly reports receiving commercial research grants from AstraZeneca and Novartis; speakers bureau honoraria from Abbott Molecular, Harrison and Star, and Qiagen; and is a consultant/advisory board member for Pfizer.

Grant Support

G.J. Riely was supported by the NIH under award number P30CA008748. C.M. Lovly was supported by the NIH under award numbers R01CA121210 and P01CA129243.

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